Introduction Mycoplasma genitalium (MG) is a sexually transmitted infection (STI) associated with urethritis in men and cervicitis and pelvic inflammatory disease (PID) in women. Azithromycin (AZ) resistance in MG, resulting in treatment failure is an issue. We compared molecular assays for detection of MG in females; a prospective cohort of adolescents with PID and a retrospective group of women at a STD clinic. We tested vaginal samples for AZ resistance with the SpeeDX assay, a real-time PCR, which detects MG and 23S rRNA AZ resistance mutations.

Methods For the prospective study, SpeeDX results were compared to 16S rRNA and pdhD real-time PCR assays, and the Aptima MG ASR assay. Samples consisted of dry transported vaginal swab samples (n=116); expressed in water. For the retrospective study we evaluated vaginal samples (n=289) stored in liquid Amies media. Nucleic acid extraction was performed utilising the Roche MagNA Pure LC robot according to manufacturer instructions. The SpeeDX assay was performed according to manufacturer instructions. For the retrospective study, results were compared to the 2 real-time PCR assays for the 16S rRNA gene and pdhD gene.

Results For the prospective study, a gold standard of 3 of 4 positive tests defined a true positive. The 16S PCR had 100% sensitivity (10/10) and 100% specificity (106/106) Kappa=1 [95% CI: 1–1], while SpeeDX PCR displayed 90% sensitivity (9/10) and 99% specificity (105/106) Kappa=0.89 [95% CI: 0.74–1]. The pdhD PCR had 100% sensitivity (10/10) and 100% specificity (106/106) Kappa=1. The Aptima MG assay had 90% sensitivity (9/10) and 94% specificity (100/106) Kappa=0.69 [0.46–0.91]. The SpeeDX 23S rRNA mutation rate was 50% (5/10). For the retrospective group, no Aptima MG was done because of transport media. A gold standard of 2 of 3 positive tests defined a true positive. The 16S PCR was 84% sensitive (21/25) and 100% specific (264/264) Kappa=0.91 [0.813–0.997]. The SpeeDX was 92% sensitive (23/25) and 99% specificity (262/264) Kappa=0.91 [0.833–0.997]. The pdhD PCR was 96% sensitive (24/25) and 99% specific (261/264) Kappa=0.91 [0.833–0.997]. The SpeeDX AZ resistance was 36% (9/25).

Conclusion: More prospective evaluation studies are required; all assays performed well. The SpeeDX PCR compared well to two real-time PCRs and the Aptima MG assay. High AZ resistance was observed. The SpeeDX assay is potentially useful for MG diagnosis and for detection of resistance to AZ